Repetitive afterglow in zirconia by pulsed near-infrared irradiation toward biological temperature sensing

Photoluminescence provides information about the surrounding environment. In this study, aiming to develop a non-invasive deep body-temperature sensing method, we investigated photoluminescence properties of afterglow zirconia (ZrO2) by pulsed near-infrared (NIR) light irradiation based on the biological temperature. Pulsed light irradiation produced optically stimulated luminescence, followed by afterglow, with the property of repeating 100 times or more. Furthermore, the basic principle of temperature measurement was demonstrated through afterglow decay curve measurements. The use of harmless ZrO2 as a sensing probe and NIR light, which is relatively permeable to living tissues, is expected to realize temperature measurements in the brain and may also facilitate optogenetic treatment.

Our research group has proposed a new temperature sensing concept for deep parts of the human body based on AG and OSL and then demonstrated a basic principle for temperature determination by the AG decay curve using a chemically-stable and harmless oxide, namely, zirconia (ZrO 2 ), as a sensing probe 9 . In addition, external laser stimulation with a wavelength (~ 650-1000 nm) in the NIR region, which has relatively high transmittance to biological tissue (i.e., biological window), enables site-selective and arbitrary-timing measurements, and we have also been able to observe OSL by irradiating a bone sample with a continuous-wave NIR-laser 9 . Meanwhile, continuous irradiation is regarded as an external source of energy in terms of temperature rise and resulting in thermal damage in biological tissue. In this study, to demonstrate biological temperature sensing using AG phosphor, we have investigated the impact of environmental temperature on the PL properties of a ZrO 2 sample under pulsed NIR irradiation. OSL, along with AG, was observed at 480 nm by repetitive pulsed irradiation, and the observation could be continued over ~ 100 cycles of pulsed irradiation. Furthermore, evaluation of temperature was also demonstrated on the basis of decay curves observed in the AG process. www.nature.com/scientificreports/ above room temperature, attributed to AG and OSL, respectively 9 . The ZrO 2 sample treated at 1400 °C for 6 h exhibits AG after ultraviolet (UV)-light exposure (254 nm), followed by OSL visually observed at the NIR-laserirradiated spot (inset). However, the presence of multiple peaks complicates the analysis of AG decay curves 9 . Therefore, prior to the TL measurement, we pre-heated the ZrO 2 sample at 323 K for 2.5 h to release electrons trapped in trapping sites, appearing below room temperature, i.e., de-trapping. According to the result, peaks A-E vanished, and only peak F at ~ 360 K, which is inactive at biological temperature, was observed in the detrapped (pre-heated) sample. Hereafter, measurements were performed using the de-trapped sample. Figure 2 shows the PL intensity of the ZrO 2 sample irradiated with repetitive pulsed NIR light at room temperature. The intensity rapidly increased immediately after irradiation and subsequently decayed (Fig. 2a). The rapid increase in PL intensity is attributed to OSL due to NIR irradiation. The moderate decay after pulsed irradiation is regarded as an AG process because the irradiation was stopped in this term. This leads us to consider that AG decay curves induced by OSL exhibit similar behavior to AG properties previously reported, i.e., the lifetime decreases as the environmental temperature of ZrO 2 increases 9,13 . Furthermore, we could confirm the rapid PL increase of over 100 times caused by repetitive irradiation (Fig. 2b). On the other hand, the maximum OSL intensity decreased over time (or repetition), e.g., I max;t~10 / I max;t~1000 ~ 0.066, Decay curve response to pulsed NIR irradiation and its temperature dependence. Next, we examined the temperature dependence on AG lifetime by pulsed irradiation. Since the release of electrons is caused by thermal excitation, the environmental temperature determines the AG lifetime (or slope of an AG curve). Assuming that AG phosphor contains only one type of electron-trapping site, the temperature dependence on the intensity of AG PL follows a simple exponential relationship: (1)  www.nature.com/scientificreports/ where I(t) represents the PL intensity, I(0) represents the initial intensity, t represents time, a represents the probability of thermal activation of a trapped electron into the conduction band, E represents the thermal activation energy of a trapped electron, s represents the frequency factor, k B represents the Boltzmann constant, and T represents temperature. We can obtain the activation energy by observing the temperature dependence on a, which corresponds to an inverse of PL lifetime ( 1/τ ). As a result, we can estimate the environmental temperature of phosphor by AG measurements. Figure 3 shows AG decay curves of ZrO 2 by single pulsed NIR irradiation at varying environmental temperatures. The decay curves observed at 313 K and 323 K seemed to be monotonically decreased, whereas the initial intensity (t < ~ 1) rapidly decayed and then decreased monotonically (Fig. 3a). This suggests that the OSL intensity partially contributed to the AG intensity; thus, we re-plotted the decay curves, which are normalized by the intensity at t = 1 s to evaluate the lifetime by fitting using a single exponential function. As a result, the de-trapped sample tended to exhibit a linear decay response with slopes that increased with increasing temperature. On the other hand, the response at 323 K deviated from the theoretical line (inset), implying that electron release from a deeper trapping site, i.e., peak F, partially contributed to AG decay curves because a trail of peak F is extended around 323 K. The evaluated lifetime and temperature could be plotted according to the Arrhenius approach, resulting in a linear relation (Fig. 3b), yielding activation energy of approximately 0.48 eV. Thus, we demonstrated that temperature can be estimated on the basis of the AG process lifetime by pulsed NIR irradiation simulations.
The activation energy in this study is comparable to that of AG in ZrO 2 previously reported 9 . To clarify the AG process in detail, we measured the PL intensity of the de-trapped ZrO 2 sample through NIR irradiation stimulation at 77 K and 323 K and compared its decay behaviors. Figure 4 depicts PL intensities detected by  www.nature.com/scientificreports/ NIR irradiation with a rectangular waveform (width: 10 s). Immediately after irradiation, obvious PL could be observed at both temperatures ( Fig. 4a). At 77 K, the PL is attributed to OSL, which is occurred by the recombination between holes and electrons released by NIR irradiation. In addition, at 323 K, the sample exhibited moderate AG decay when the irradiation was stopped, whereas a rapid decrease in the intensity was observed at 77 K, indicating that AG is barely observed at 77 K. These measurements lead to the conclusion that, at 323 K, electrons trapped in trapping sites, which are inactive at biological temperature (peak F), are released by NIR irradiation, and subsequently, these electrons are partially re-trapped at the thermally active site (peak E) via transition to the conduction band. Eventually, the AG process is observed (route A; Fig. 4b). At 77 K, although electrons released by NIR irradiation are trapped by active sites as well, the thermal release is suppressed because this temperature is significantly lower than peak E and others (route B). According to the result, only OSL can be observed. Therefore, it is reasonable to assume that AG observed immediately after NIR irradiation is regarded as typical AG, and consequently, they are identical. Although the origin of AG in ZrO 2 is still controversial, Iwasaki et al. suggest that the trapping sites in AG was closely related to the presence of F + -center (or V o · in Kröger-Vink notation) on the basis of measurement of electron spin resonance 13 .
This study can demonstrate the availability of pulsed NIR irradiation, but we should also consider converting the PL wavelength to be compatible with the biological window range. The conversion of PL to a longer wavelength in ZrO 2 using an appropriate dopant has been reported 14,15 , and a similar study is underway.

Summary
The temperature dependence of PL properties of ZrO 2 under pulsed NIR-laser irradiation was investigated for biological temperature sensing applications by decay curve measurements. We demonstrated that environmental temperatures can be estimated from the AG lifetime through the Arrhenius approach. The use of low-toxicity ZrO 2 as a sensing probe is expected to reduce stress in living organisms. Repetitive pulsed NIR-laser irradiation is expected to not only reduce damage to biological tissue but also shorten the time and improve the accuracy of repetitive temperature measurements.
Non-invasive therapies based on optogenetics have been reported to control memory and sociality via light irradiation of local areas of the brain and neurons, but in these experiments, optical fibers/LEDs are inserted directly into the irradiated areas of mice [16][17][18][19] . Therefore, AG and OSL caused by external pulsed NIR irradiation using AG probes may contribute to optogenetic therapy by enabling non-invasive and site-selective optical simulations of local areas.

Methods
We prepared the ZrO 2 sample by annealing oxygen-defect activation, and the sample exhibited a broad PL peak at ~ 480 nm due to AG and OSL 9,13 . Therefore, in this study, we also prepared another ZrO 2 sample by referring to the previous study: Commercial reagent-grade ZrO 2 powder (purity: 99.9%, Soekawa Chem. Co., Ltd.) was thermally treated at 1400 °C for 6 h to introduce oxygen defects. The heating rate was 10 K/min, and the target temperature was maintained for 6 h in an electric furnace under atmospheric conditions. After the treatment, the ZrO 2 powder was cooled in the furnace, and finally, the ZrO 2 sample was obtained.
At room temperature, ZrO 2 crystallizes in monoclinic system, and undergoes a phase-transition to tetragonal system at ~ 1170 °C. Thermal treatment above the phase-transition temperature introduces oxygen defects, which are associated with the phase transition 13 . Since tetragonal ZrO 2 has been found to show no PL 20 , structural analysis of the obtained sample was performed by means of an X-ray diffraction (XRD; Cu-Kα). As the result, the phase of treated ZrO 2 was identified to be monoclinic (Fig. 5a). In addition, a transmission electron microscopy (TEM) revealed clear lattice pattern in the ZrO 2 grain, suggesting the preservation of crystallinity (Fig. 5b). www.nature.com/scientificreports/ The PL intensity as a function of time (decay curve) was measured using a spectrofluorometer with a xenon lamp as the excitation source. The environmental temperature of the ZrO 2 sample during decay curve measurement was controlled using a cryostat. The PL intensity as a function of temperature (TL spectrum) was also measured using the spectrofluorometer conjugated with the cryostat. The intensity was detected at 480 nm in the decay curve during TL measurements with a heating rate of 1 K/min. In addition, prior to these measurements, the ZrO 2 sample was excited at 280 nm, corresponding to the PL excitation peak, for 1 h. The wavelengths for PL and PL excitation were selected according to a previous study 9 . An NIR-laser diode (808 nm, 90 mW) was installed on the cryostat, and pulsed light was generated by combining a function generator with the power supply unit. The waveform was a square wave, with a pulse width of 0.5 s.

Data avairability
The data and material used and/or analysed during this study are available from the corresponding author on reasonable request.